Cultivation of novel <em>Atribacterota</em> from oil well provides new insight into their diversity, ecology, and evolution in anoxic, carbon-rich environments

Background: The Atribacterota are widely distributed in the subsurface biosphere. Recently, the first Atribacterota isolate was described and the number of Atribacterota genome sequences retrieved from environmental samples has increased significantly; however, their diversity, physiology, ecology, and evolution remain poorly understood. Results: We report the isolation of the second member of Atribacterota, Thermatribacter velox gen. nov., sp. nov., within a new family Thermatribacteraceae fam. nov., and the short-term laboratory cultivation of a member of the JS1 lineage, Phoenicimicrobium oleiphilum HX-OS.bin.34^TS, both from a terrestrial oil reservoir. Physiological and metatranscriptomics analyses showed that Thermatribacter velox B11^T and Phoenicimicrobium oleiphilum HX-OS.bin.34^TS ferment sugars and n-alkanes, respectively, producing H₂, CO₂, and acetate as common products. Comparative genomics showed that all members of the Atribacterota lack a complete Wood-Ljungdahl Pathway (WLP), but that the Reductive Glycine Pathway (RGP) is widespread, indicating that the RGP, rather than WLP, is a central hub in Atribacterota metabolism. Ancestral character state reconstructions and phylogenetic analyses showed that key genes encoding the RGP (fdhA, fhs, folD, glyA, gcvT, gcvPAB, pdhD) and other central functions were gained independently in the two classes, Atribacteria (OP9) and Phoenicimicrobiia (JS1), after which they were inherited vertically; these genes included fumarate-adding enzymes (faeA; Phoenicimicrobiia only), the CODH/ACS complex (acsABCDE), and diverse hydrogenases (NiFe group 3b, 4b and FeFe group A3, C). Finally, we present genome-resolved community metabolic models showing the central roles of Atribacteria (OP9) and Phoenicimicrobiia (JS1) in acetate- and hydrocarbon-rich environments. Conclusion: Our findings expand the knowledge of the diversity, physiology, ecology, and evolution of the phylu. Authors: Jiao, Jian-Yu [Sun Yat-Sen Univ., Guangzhou (China)]; Ma, Shi-Chun [Biogas Institute of Ministry of Agriculture and Rural Affairs, Chengdu (China)]; Salam, Nimaichand [Sun Yat-Sen Univ., Guangzhou (China); National Agri-Food Biotechnology Institute, Mohali (India)]; Zhou, Zhuo [Biogas Institute of Ministry of Agriculture and Rural Affairs, Chengdu (China)]; Lian, Zheng-Han [Sun Yat-Sen Univ., Guangzhou (China)]. DOE Contract: AC02-05CH11231. Subjects: 59 BASIC BIOLOGICAL SCIENCES; Atribacteria; Atribacterota; Carbohydrate fermentation; Enrichment; Hydrocarbon degradation; Oil reservoir; Phoenicimicrobiia; Pure culture; Reductive glycine pathway; Wood-Ljungdahl pathway